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为了获得与盐碱胁迫相关的14-3-3蛋白基因的表达模式,基于羊草14-3-3蛋白的EST序列,通过RACE方法从羊草中克隆得到的Lc14-3-3基因,并利用生物信息学分析获得了其基因信息,利用荧光定量PCR(qRT-PCR)分析获得了羊草植株在Na_2CO_3胁迫下Lc14-3-3基因的表达模式。结果表明:Lc14-3-3基因编码区全长为786 bp,编码261个氨基酸,推测蛋白质分子量为29.26 k D,理论等电点(p I)为4.83,与已知的单子叶植物来源的同类基因同源性较高,相似性为88%~100%。q RT-PCR表达模式分析表明,羊草14-3-3基因在无胁迫处理情况下有本底水平表达。在200 mmol/L的Na_2CO_3胁迫不同时间后,羊草叶片中Lc14-3-3基因的表达量在6~24 h之间表达量明显上升,但胁迫持续48 h时,Lc14-3-3表达量降至较低水平。但随着胁迫时间增加Lc14-3-3表达量骤增,96 h后高于对照组10倍左右。羊草根部的Lc14-3-3基因表达量在胁迫处理后6~24 h间为无胁迫条件的2倍,在胁迫处理48 h后下降,随着胁迫时间增加,96 h表达量为对照组11倍左右。此结果为进一步研究抗盐碱基因提供理论基础。
In order to obtain the expression pattern of 14-3-3 protein gene related to salt-alkali stress, the Lc14-3-3 gene was cloned from Leymus chinensis by RACE based on the EST sequence of Leymus chinensis 14-3-3 protein. The gene information was obtained by bioinformatics analysis, and the expression pattern of Lc14-3-3 gene in Leymus chinensis plants under Na_2CO_3 stress was obtained by qRT-PCR. The results showed that the full-length coding region of Lc14-3-3 gene was 786 bp, encoding 261 amino acids. The predicted molecular weight of Lc14-3-3 gene was 29.26 kD and the theoretical isoelectric point (pI) was 4.83. Compared with the known monocotyledonous High homology of similar genes, similarities of 88% to 100%. q RT-PCR expression pattern analysis showed that the 14-3-3 gene of Leymus chinensis had a background level under the condition of no stress. After 200 mmol / L Na_2CO_3 stress, the expression of Lc14-3-3 gene in Leymus chinensis leaves increased significantly from 6 to 24 h, but the expression of Lc14-3-3 was up to 48 h The amount dropped to a lower level. However, with the increase of stress time, the expression of Lc14-3-3 increased sharply and was about 10 times higher than that of the control group after 96 hours. The expression level of Lc14-3-3 in the roots of Leymus chinensis was twice as high as that of the non-stress conditions 6-24 h after stress, and decreased after 48 h of stress treatment. The expression of Lc14-3-3 at 96 h was the control group About 11 times. The results provide a theoretical basis for further study of salt-resistant genes.